Periodic Reporting for period 1 - VICI-DISEASE (PROTECTION FROM VIRUSES WITH EPIDEMIC AND PANDEMIC DISEASE OUTBREAK POTENTIAL THROUGH DEVELOPMENT AND CLINICAL TESTING OF A NOVEL CAPSID VIRUS LIKE PARTICLE (CVLP) VACCINE)
Periodo di rendicontazione: 2023-12-01 al 2025-05-31
The major challenge for the modern vaccines is the induction of long-term protective immunity, as clearly demonstrated by fast waning of protection for current COVID-19 vaccines. VICI-DISEASE is an ambitious project which combines existing cutting-edge expertise in a tried and tested consortium, with new advances in this critical field. The consortium’s main objective is to develop a vaccine candidate portfolio and perform a clinical proof-of-concept study, to enable stocks of vaccine candidates ready for further development (Phase 2&3) in case of pandemic outbreaks. The primary candidate will be Nipah virus (NiV), a virus leading to high-mortality disease with no vaccines or treatments available. Our vaccine is based on NiV G protein displayed on capsid Virus Like Particle (cVLP), enabled by adapting template processes from our recent COVID-19 vaccine (currently in Phase 3), and expected to provide best-in-class longevity as shown for COVID-19. This cVLP vaccine will be uniquely positioned to help prevent future NiV epidemics and pandemics.
The VICI-DISEASE consortium has the objective to develop and perform a clinical proof-of-concept study for a novel NiV vaccine (G protein displayed on cVLP), which is:
• highly effective (>90% protection),
• long-term protection (>2 years),
• rapidly acting (within 2 weeks),
• utilising cVLP technology which is the only clinically proven vaccine platform capable of generating high-level neutralizing antibody responses without the need for adjuvant,
• by adapting template processes established for our COVID-19 vaccine (currently in Phase 3),
• and tested within 48 months in a Phase 1/2a clinical study,
• to help protect medical workers and the public from future NiV and Hendra virus (HeV) outbreaks,
• and establish a pipeline of novel filovirus vaccines through pre-clinical proof-of-concept studies.
The VICI-DISEASE consortium has the objective to develop and perform a clinical proof-of-concept study for a novel NiV vaccine (G protein displayed on cVLP), which is:
• highly effective (>90% protection),
• long-term protection (>2 years),
• rapidly acting (within 2 weeks),
• utilising cVLP technology which is the only clinically proven vaccine platform capable of generating high-level neutralizing antibody responses without the need for adjuvant,
• by adapting template processes established for our COVID-19 vaccine (currently in Phase 3),
• and tested within 48 months in a Phase 1/2a clinical study,
• to help protect medical workers and the public from future NiV and Hendra virus (HeV) outbreaks,
• and establish a pipeline of novel filovirus vaccines through pre-clinical proof-of-concept studies.
A broad panel of vaccine antigen candidates was evaluated for recombinant expression in both HEK cells (by UCPH) and Drosophila S2 cells (by Expres2ion). Early-stage efforts focused on the Nipah virus G and F proteins, as well as G proteins from related strains (Bangladesh, Malaysia) and other henipaviruses (Hendra, Ghana, Langya). In total, approximately 100 antigen constructs were generated, each genetically fused to either a Tag or Catcher domain to facilitate site-specific conjugation to a capsid Virus-Like Particle (cVLP). These constructs were tested across a range of cVLP-display strategies—including multivalent antigen designs—and within both protein- and mRNA-based vaccine platforms. Multiple linker configurations were also assessed to optimize expression, cVLP coupling efficiency, and antigen stability.
Recombinant expression in S2 cells yielded high levels of non-aggregated antigens that retained correct conformation, as confirmed by binding to conformation-sensitive monoclonal antibodies. These antigens could be efficiently coupled at high density to the AP205 cVLP, forming stable antigen:cVLP complexes. Selected lead candidates were subsequently evaluated for immunogenicity in mice, where they induced strong ELISA-binding and virus-neutralizing antibody responses.
In parallel, a genetic vaccine platform was used to identify antigen candidates suitable for either protein- or mRNA-based vaccine development. This approach supported the selection of the protein vaccine lead and served as a mitigation strategy by enabling identification of promising mRNA vaccine candidates. Antigens were initially screened for expression in HEK cells and evaluated for their ability to form secreted antigen:cVLP complexes. Promising designs were evaluated for immunogenicity in mice. The mRNA vaccines induced strong antigen-specific ELISA-binding titers as well as high virus-neutralizing responses.
Direct comparison of selected constructs delivered as mRNA or protein-based cVLP vaccines demonstrated that both vaccine platforms succeeded in inducing robust ELISA antibody titers, comparable to those measured in mice vaccinated with a control NiV vaccine, similarly to mice surviving a lethal dose of NiV challenge. These antibody levels also correlated with potent neutralization of Nipah virus, surpassing titers observed in convalescent human sera (WHO standard) and benchmark vaccine candidates.
Furthermore, cellular immune responses were evaluated by measuring antigen-specific CD8⁺ T cell responses in the spleens of immunized mice. Here, only the mRNA-based vaccines elicited robust CD8⁺ T cell responses, which were not observed following immunization with protein-based cVLP vaccines.
Recombinant expression in S2 cells yielded high levels of non-aggregated antigens that retained correct conformation, as confirmed by binding to conformation-sensitive monoclonal antibodies. These antigens could be efficiently coupled at high density to the AP205 cVLP, forming stable antigen:cVLP complexes. Selected lead candidates were subsequently evaluated for immunogenicity in mice, where they induced strong ELISA-binding and virus-neutralizing antibody responses.
In parallel, a genetic vaccine platform was used to identify antigen candidates suitable for either protein- or mRNA-based vaccine development. This approach supported the selection of the protein vaccine lead and served as a mitigation strategy by enabling identification of promising mRNA vaccine candidates. Antigens were initially screened for expression in HEK cells and evaluated for their ability to form secreted antigen:cVLP complexes. Promising designs were evaluated for immunogenicity in mice. The mRNA vaccines induced strong antigen-specific ELISA-binding titers as well as high virus-neutralizing responses.
Direct comparison of selected constructs delivered as mRNA or protein-based cVLP vaccines demonstrated that both vaccine platforms succeeded in inducing robust ELISA antibody titers, comparable to those measured in mice vaccinated with a control NiV vaccine, similarly to mice surviving a lethal dose of NiV challenge. These antibody levels also correlated with potent neutralization of Nipah virus, surpassing titers observed in convalescent human sera (WHO standard) and benchmark vaccine candidates.
Furthermore, cellular immune responses were evaluated by measuring antigen-specific CD8⁺ T cell responses in the spleens of immunized mice. Here, only the mRNA-based vaccines elicited robust CD8⁺ T cell responses, which were not observed following immunization with protein-based cVLP vaccines.
- In vitro POC for a mRNA and protein cVLP. The new generation mRNA vaccine has been developed throughout the exploration of a NiV vaccine. The work carried out on the mRNA vaccine encoding for a Tag/Catcher cVLP technology shows that it is possible to express the tag/catcher fused to the antigen and to cVLP from mRNA in vitro leading to the secretion of NiVG decorated cVLP. The mRNA vaccine encoding for a tag/catcher VLP technology is really promising in the context of a NiV vaccine as it induces high antibody titers with high neutralizing capacity. This new technology is also promising for the general vaccine field as it would combine the advantages of mRNA vaccine (rapid production, high immunogenicity) and VLP vaccines (high immunogenicity, long-lasting protection, dose-sparring potential).
- Our current G-head vaccines demonstrate in vitro neutralization 13-fold higher than the human WHO standard in mice for the mRNA vaccine and 9-fold for the protein vaccine. Thus, our vaccines are inducing stronger antibody responses compared to human surviving a NiV infection (WHO standard).
- Discovery of dual display on the VLP: new antigen designs have been explored, where 2 antigens can be closely presented by having a double display (same protein for homo-display or 2 different proteins for hetero-display). This high-density presentation of antigens may improve the induction of cross-reactive immune responses against various henipaviruses by directing the immune response toward conserved epitopes. Although this design is extremely relevant for NiV vaccine development, this method could also be utilized in the context of many other vaccines that require the generation of broadly reactive antibodies.
- New protein designs for vaccines from Leipzig's partners (ULEI): ULEI has produced new NiV antigen designs that are currently explored within this consortium. ULEI uses a vaccine design pipeline that uses artificial intelligence (AI) to introduce stabilizing mutations. More specifically, an interface with the bioinformatic software suite Rosetta with AI technologies, that was recently developed at ULEI (Ertelt et al. Science Advances), is used to sample mutations from different strategies, such as evolution-derived, symmetry aware, from graph neural networks, or large language models. By using these methods, new stabilized proteins have been established, which would be particularly useful for the NiV fusion protein (F protein) which is inherently hard to produce, but might be required for long-lasting and broad protection. These methods can be further applied for any protein used in vaccine designs, beyond this consortium.
- Our current G-head vaccines demonstrate in vitro neutralization 13-fold higher than the human WHO standard in mice for the mRNA vaccine and 9-fold for the protein vaccine. Thus, our vaccines are inducing stronger antibody responses compared to human surviving a NiV infection (WHO standard).
- Discovery of dual display on the VLP: new antigen designs have been explored, where 2 antigens can be closely presented by having a double display (same protein for homo-display or 2 different proteins for hetero-display). This high-density presentation of antigens may improve the induction of cross-reactive immune responses against various henipaviruses by directing the immune response toward conserved epitopes. Although this design is extremely relevant for NiV vaccine development, this method could also be utilized in the context of many other vaccines that require the generation of broadly reactive antibodies.
- New protein designs for vaccines from Leipzig's partners (ULEI): ULEI has produced new NiV antigen designs that are currently explored within this consortium. ULEI uses a vaccine design pipeline that uses artificial intelligence (AI) to introduce stabilizing mutations. More specifically, an interface with the bioinformatic software suite Rosetta with AI technologies, that was recently developed at ULEI (Ertelt et al. Science Advances), is used to sample mutations from different strategies, such as evolution-derived, symmetry aware, from graph neural networks, or large language models. By using these methods, new stabilized proteins have been established, which would be particularly useful for the NiV fusion protein (F protein) which is inherently hard to produce, but might be required for long-lasting and broad protection. These methods can be further applied for any protein used in vaccine designs, beyond this consortium.